P706: RESISTANCE TO HYPOMETHYLATING AGENTS (HMA) IN HIGH-RISKMYELODYSPLASTIC SYNDROME (HR-MDS): THE ROLE OF THE ADENOSINEDEAMINASE ACTING ON RNA 1 (ADAR1) ENZYME
Authors: Despoina Dimitriou, Eleftheria Lamprianidou, Athanasios Tasis, Theodoros Spiropoulos, Chrysa Kimparidou, Stavros Papadakis, Konstantinos Liapis, Menelaos Papoutselis, George Vrachiolias, Ioannis Mitroulis, Ioannis Kotsianidis
Journal: HemaSphere (2023)

Background: The HMAs azacytidine (AZA) and decitabine (DAC) are the mainstay of treatment for HR-MDS, but their exact mechanism of action remains largely unknown. Though HMAs induce a viral mimicry response on cancer lines, the latter was not associated with clinical responses when measured in primary leukemia cells. The differential expression of the A-to-I RNA editing enzyme, ADAR1, which depletes the immunogenic dsRNAs may be responsible for the disconnection between the induction of viral mimicry and clinical response to HMA. Aims: We sought to investigate the role of ADAR1 in the HMA resistance in HR-MDS patients utilizing bone marrow samples from well-annotated patient cohorts. Methods: We measured ADAR1 levels and its two isoforms, ADAR1p150 and ADAR1p110, by quantitative real-time PCR in purified primary CD34+ cell subsets. We also performed functional immunophenotyping of STAT3 phosphorylation in primary CD34+ cells. Results: Non-responders to HMAs (N=25, AZA=15, DAC=10) demonstrated significantly higher pretreatment ADAR1 levels compared to responders (N=22, AZA=18, DAC=4, p=0.0047), but the difference was significant only in ΑΖΑ-treated patients (p=0.0006, Figure 1a). We further investigated the expression of the interferon-inducible ADAR1p150 isoform and the constitutively expressed ADAR1p110 one. The expression levels of both isoforms were higher in non-responders compared to responders to AZA (pp150 AZA=0.0080, pp110 AZA= 0.0066, Figures 1b-c). The level of cellular differentiation where AZA acts by inducing the viral mimicry state is still unidentified; therefore, we measured ADAR1 levels in FACS-sorted hematopoietic stem and progenitor cell subpopulations. We observed significantly higher ADAR1p150 levels in the more primitive Lin-CD34+CD38- compartment of non-responders compared to responders to AZA (p= 0.0038, Figure 1d), but not in the Lin-CD34+CD38+ committed progenitors (p= 0.9879) or the erythroblasts (p=0.4079). Recent data support the existence of a positive feedback regulatory loop between STAT3b and ADAR1, in which STAT3b promotes ADAR1 expression while, in turn, ADAR1 contributes to the stabilization of STAT3b. We studied the expression of STAT3a and STAT3b in purified total CD34+ cells from HMA-treated HR-MDS patients, but we observed no significant associations with the response to HMAs (pSTAT3a = 0.3277, pSTAT3b = 0.2203, Figures 1e-f). However, we found a positive correlation between the relative expression of STAT3b and ADAR1p150 isoform only in non-responders to AZA patients (p=0.008). We further correlated the STAT3 phosphorylation levels of CD34+ progenitors with ADAR1 levels of the same cells. Both total ADAR1 and the p150 isoform were positively correlated with the G-CSF-inducible STAT3 levels in non-responders (pADAR1 = 0.011, pp150 = 0.025). Summary/Conclusion: Our findings implicate the ADAR1 pathway in the resistance to AZA and further corroborate the potential existence of a vicious cycle among STAT3 and ADAR1 in the leukemic progenitors of HR-MDS patients. If confirmed in larger cohorts, our data argue for the utilization of ADAR1 as both a therapeutic target and a predictor of AZA response. Acknowledgments: “InTechThrace: Integrated Technologies in biomedical research: multilevel biomarker analysis in Thrace”, MIS 5047285.Keywords: Myelodysplastic syndrome, Hypomethylating agents, STAT3, Epigenetic